A large number of pathological conditions in humans, animals, and plants are now understood at the genetic level. With the announced completion of the mapping of the human genome, it is expected that the genetic basis of many more human diseases will be identified in the coming years. Analysis of DNA from an individual, therefore, can, in principle, allow genetically based conditions to be diagnosed or to be identified in the absence of overt symptoms. This is advantageous for many conditions such as metabolic disorders in which early diagnosis can prevent serious medical complications later in life.
Methods of analyzing DNA sequences, which are often referred to generically as genotyping, are known in the art. In very general terms, to determine whether the DNA in a sample corresponds to a particular disease condition whose genetic sequence is known, the sample is exposed to nucleic acid probes associated with that disease, under conditions that allow hybridization. The nucleic acid probes are labeled making it possible to detect whether the probes have hybridized to the DNA sample. In one technique, the probes are arranged in arrays on chips, with each probe assigned to a specific location. After exposing the array to a labeled DNA sample, scanning devices can examine each location in the array and determine whether a target molecule has interacted with the probe at that location. Array chips are provided commercially, for example, by Affymetrix (Santa Clara, Calif.) and are described in patents assigned to Affymetrix (See, for example U.S. Pat. Nos. 6,045,996, 5,858,659, and 5,925, 525, and references therein.). Arrays have also been used for DNA sequencing applications such as the Sequencing by Hybridization approaches described, for example, in U.S. Pat. Nos. 6,025,136, 6,018,041, 5,525,464, and 5,202, 231.
While methods of genotyping for disease diagnostics are available, in order for the methods to be useful in a public health setting, they need to be reasonable in cost. For example, although relevant genetic assays are known, neonatal screening is currently done by mass spectrometric methods primarily because of cost considerations. Secondly, DNA diagnostics in a public health setting need to be practical for application to multiple samples and to genetic conditions in which mass spectrometric methods are difficult or intractable. The requirement of multiple samples may be addressed by using multiple array chips, which are processed simultaneously. As described in U.S. Pat. No. 5,545,531 to Rava et al., a format including a standard 96-well microtiter plate containing an array chip at the bottom of each well can be used. To perform the same test on many patient samples, each patient sample, in solution, is labeled and introduced into a different well, each of which has an identical array chip. Thus, in this method, a separate array chip is used for each sample, which may be costly for widespread use because of the fixed per-patient costs of arrays, reagents, sample processing, and so forth.
U.S. Pat. No. 5,807,522 to Brown et al. describes a method of screening multiple patients against known mutations in a disease gene using multiple microarrays of patient genomic DNA and probe DNA fragments representing all known mutations of a given gene. The microarrays are fabricated on a sheet of plastic-backed nitrocellulose with silicone rubber barrier elements between individual arrays to prevent cross contamination. All microarrays are processed as a single sheet of material. However, the method of Brown et al. uses a separate microarray for each mutated allele or genetic marker screened.
Thus, there is a need for a method of genotyping with sufficient precision for diagnostic use, that is affordable and that provides sufficient throughput for large-scale use. Ideally, such a method would allow multiple patients to be screened for multiple diseases in a single assay. More generally, the method would allow multiple samples from any source of human, animal, plant, or microbial material to be screened for alleles at multiple genetic loci in a single assay.